CN112816410A - Depth imaging method and system of TIRF illumination - Google Patents
Depth imaging method and system of TIRF illumination Download PDFInfo
- Publication number
- CN112816410A CN112816410A CN202011637113.4A CN202011637113A CN112816410A CN 112816410 A CN112816410 A CN 112816410A CN 202011637113 A CN202011637113 A CN 202011637113A CN 112816410 A CN112816410 A CN 112816410A
- Authority
- CN
- China
- Prior art keywords
- imaged
- depth
- sample
- tirf
- imaging
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000003384 imaging method Methods 0.000 title claims abstract description 77
- 238000005286 illumination Methods 0.000 title claims abstract description 63
- 230000005540 biological transmission Effects 0.000 claims abstract description 39
- 238000000034 method Methods 0.000 claims abstract description 10
- 239000000835 fiber Substances 0.000 claims description 10
- 239000011521 glass Substances 0.000 claims description 10
- 238000007654 immersion Methods 0.000 claims description 8
- 238000000799 fluorescence microscopy Methods 0.000 claims description 6
- 239000013307 optical fiber Substances 0.000 claims description 2
- 238000010586 diagram Methods 0.000 description 4
- 239000006059 cover glass Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000035515 penetration Effects 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/01—Arrangements or apparatus for facilitating the optical investigation
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6402—Atomic fluorescence; Laser induced fluorescence
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Immunology (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Pathology (AREA)
- General Physics & Mathematics (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Optics & Photonics (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
- Microscoopes, Condenser (AREA)
Abstract
The invention discloses a depth imaging method and a depth imaging system of TIRF illumination, wherein the method comprises the following steps: during TIRF illumination, incident light is totally reflected in front of a sample to be imaged, an evanescent wave is generated, and the generated evanescent wave is transmitted to the sample to be imaged; changing the incident angle of incident light to change the transmission depth of evanescent waves; when evanescent waves with different transmission depths respectively irradiate the samples to be imaged, the STORM imaging unit images the samples to be imaged, all the images of the samples to be imaged are calculated through a preset image reconstruction algorithm, the images of the samples to be imaged with different depths are obtained, and depth imaging is achieved. The invention realizes imaging in a certain depth direction through different TIRF illumination based on a STORM + TIRF mode.
Description
Technical Field
The invention relates to the technical field of microscopic imaging, in particular to a depth imaging method and system of TIRF illumination.
Background
STORM is in the field of microscopic imaging and is based on the principle that when light is incident on cells with fluorescent stains, the cells emit random fluorescent scintillation dots. Through multiple imaging, the fluorescence scintillation point at each position can be finally obtained, and the position of the scintillation point is the position point of the cell. When light irradiates on cells, because of random scintillation of fluorescence, after one-time scintillation, the position of each scintillation point can be finally obtained, and then images of different positions of the cells can be obtained. When the fluorescent cell is imaged, the scintillation point is reconstructed by a picture of fluorescent molecules which randomly flicker for a period of time, so that super-resolution imaging is realized.
TIRF illumination is a total reflection and generation of evanescent waves, i.e. light is reflected internally in an optically dense medium at total reflection, and according to the physical optics section, light generates a thin layer of evanescent waves in an optically thinner medium. Typical penetration under effective illumination is only 50nm to 100nm, only fluorescent molecules near the cover glass surface (approach) can be excited, and far-field molecules are not excited.
The STORM microscope imaging adopts evanescent waves generated by TIRF illumination to irradiate cells with fluorescent staining, and an illumination area can be thinner during the TIRF illumination, so that stray light during imaging is reduced, and the signal to noise ratio of imaging is improved. However, the combination of STORM + TIRF does not allow depth imaging because the evanescent wave travels only in a thin layer and only against an optically thinner medium near the optically denser medium.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides a depth imaging method and a depth imaging system of TIRF illumination capable of realizing depth imaging.
The purpose of the invention is realized by the following technical scheme:
a method of depth imaging of TIRF illumination, comprising: during TIRF illumination, incident light is totally reflected in front of a sample to be imaged, an evanescent wave is generated, and the generated evanescent wave is transmitted to the sample to be imaged; changing the incident angle of incident light to change the transmission depth of evanescent waves; when evanescent waves with different transmission depths respectively irradiate the samples to be imaged, the STORM imaging unit images the samples to be imaged, all the images of the samples to be imaged are calculated through a preset image reconstruction algorithm, the images of the samples to be imaged with different depths are obtained, and depth imaging is achieved.
Preferably, a method of depth imaging with TIRF illumination comprises:
s1, the incident angle of the incident light is theta during TIRF illumination1Transmission depth of H1Fluorescence imaging by STORM as S1；
S2, changing the incident angle to theta2At the time of the depth of transmission of H2Fluorescence imaging by STORM as S2(ii) a Wherein H2>H1；
S3, calculating the transmission depth H1-H2The corresponding image of the sample to be imaged is S2-S1；
S4, repeatedly changing different incidence angles until obtaining the transmission depth Hn-Hn-1Corresponding sample image S to be imagedn-Sn-1And imaging of samples to be imaged at different depths is realized.
Preferably, the incident angle of the incident light and the transmission depth of the evanescent wave are expressed as:
where θ is the incident angle of the incident light, d is the transmission depth of the evanescent wave, and n1Is the refractive index, n, of the objective lens in front of the sample to be imaged in TIRF illumination2Is the refractive index of the sample to be imaged.
Preferably, a method of depth imaging with TIRF illumination further comprises: intensity compensation is performed for evanescent wave illumination at different depths to correct for imaging intensity variations due to evanescent wave depth.
A TIRF illuminated depth imaging system, comprising: a STORM imaging unit and a TIRF illumination unit; the TIRF illuminating units respectively generate evanescent waves with different transmission depths to irradiate the samples to be imaged, and the STORM imaging units all image the samples to be imaged; the sample to be imaged is clamped between an upper glass slide and a lower glass slide, immersion oil is filled between the lower glass slide and the objective lens, and the refractive indexes of the lower glass slide, the immersion oil and the objective lens are the same and are all larger than the refractive index of the sample to be imaged.
Preferably, the TIRF lighting unit comprises: an illumination light source, a middle fold-back mirror and an objective lens; laser emitted by the illumination light source is reflected by the intermediate turning mirror, is transmitted by the objective lens and then irradiates the lower glass slide for total reflection, and evanescent waves are generated and irradiate the sample to be imaged.
Preferably, the illumination light source includes: a fiber light source and a collimating lens group; laser emitted by the optical fiber light source passes through the collimating lens group and then irradiates the middle turning mirror.
Preferably, the TIRF illumination unit generates evanescent waves of different transmission depths respectively including: the incidence angle of the sample to be imaged is changed by changing the movable piece of the TIRF lighting unit, and a relation curve of the moving position and the incidence angle is obtained by measuring the relation between the moving position of the movable piece and the incidence angle of the sample to be imaged at the moment.
Preferably, the TIRF illumination units respectively generate evanescent waves with different transmission depths to irradiate onto the sample to be imaged, and the STORM imaging units each image the sample to be imaged, including:
the method comprises the following steps: moving the movable piece to a position L0, wherein the depth of the evanescent wave is d0, the STORM imaging camera focuses to d0 for clear imaging, and the imaging of the sample to be imaged is S0；
Step two: moving the movable piece to L1, wherein the corresponding depth of the evanescent wave is d0 +. DELTA.d, the depth of the evanescent wave is d1 ═ d0 +. DELTA.d, and the sample to be imaged is imaged as S1(ii) a Wherein, the delta d is an equal division interval after N equal division is carried out on the transmission depth of the evanescent wave;
step three: moving the movable piece to L2, wherein the corresponding depth of evanescent wave is d0+2 Δ d, and the sample to be imaged is imaged as S3；
Step four: continuously moving the movable piece until the position of the movable piece is LmMoving the objective lens in front of the sample to be imaged by a distance m delta d/n1 when the depth of the evanescent wave is d0+ m delta d, wherein the depth of the evanescent wave is d0+ m delta d, and the sample to be imaged is imaged as Sm；
Step five: repeating the second step to the fourth step, moving the movable piece until the position of the movable piece is LNWhen the sample to be imaged is imaged as SN；
Step six: and sequentially subtracting the images of two adjacent samples to be imaged to obtain delta S1, delta S2 and … delta SN respectively, wherein the images obtained by subtracting are the images of the samples to be imaged at different depths.
Preferably, the mobile element comprises: any one of a collimating lens group, a fiber optic light source, and an illumination light source.
Compared with the prior art, the invention has the following advantages:
according to the invention, the transmission depth of evanescent waves is changed by changing the incident angle of incident light during TIRF illumination; when evanescent waves with different transmission depths respectively irradiate the samples to be imaged, the STORM imaging unit images the samples to be imaged, all the images of the samples to be imaged are calculated through a preset image reconstruction algorithm, the images of the samples to be imaged with different depths are obtained, and depth imaging is achieved. Therefore, the invention realizes imaging in a certain depth direction through different TIRF illumination based on the STORM + TIRF mode.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this application, illustrate embodiments of the invention and, together with the description, serve to explain the invention and not to limit the invention. In the drawings:
FIG. 1 is a flow chart illustrating the TIRF illuminated depth imaging method of the present invention.
FIG. 2 is a graph of incident angle of incident light versus transmission depth of evanescent waves in accordance with the present invention.
FIG. 3 is a graph of depth intensity versus illumination depth for the present invention.
FIG. 4 is a diagram showing the relationship between the total reflection angle and the maximum aperture angle of the objective lens, immersion oil and a sample to be imaged.
FIG. 5 is a schematic diagram of the TIRF illuminated depth imaging system moving the collimating lens group of the present invention.
FIG. 6 is a schematic diagram of a TIRF illuminated depth imaging system fiber optic source of the present invention.
FIG. 7 is a schematic diagram of the TIRF illuminated depth imaging system moving illumination sources of the present invention.
Detailed Description
The invention is further illustrated by the following figures and examples.
A method of depth imaging of TIRF illumination, comprising: during TIRF illumination, incident light is totally reflected in front of a sample to be imaged, an evanescent wave is generated, and the generated evanescent wave is transmitted to the sample 1013 to be imaged; changing the incident angle of incident light to change the transmission depth of evanescent waves; when evanescent waves with different transmission depths are transmitted to the sample 1013 to be imaged respectively, the STORM imaging unit images the sample 1013 to be imaged, all images of the sample 1013 to be imaged are calculated through a preset image reconstruction algorithm, imaging images of the sample 1013 to be imaged with different depths are obtained, and depth imaging is achieved.
It should be noted that the preset image reconstruction algorithm is the prior art and is not an improvement point of the present invention. And the STORM principle is also prior art. The sample 1013 to be imaged is a cell.
Referring to fig. 1, a depth imaging method using TIRF illumination includes:
s1, the incident angle of the incident light is theta during TIRF illumination1Transmission depth of H1Fluorescence imaging by STORM as S1；
S2, changing the incident angle to theta2At the time of the depth of transmission of H2Fluorescence imaging by STORM as S2(ii) a Wherein H2>H1；
S3, calculating the transmission depth H1-H2The corresponding image of the sample 1013 to be imaged is S2-S1；
S4, repeatedly changing different incidence angles until obtaining the transmission depth Hn-Hn-1Corresponding image S of sample 1013 to be imagedn-Sn-1And imaging of samples 1013 to be imaged at different depths is achieved.
Referring to fig. 2, the incident angle of incident light and the transmission depth of the evanescent wave are expressed as:
where θ is the incident angle of the incident light, d is the transmission depth of the evanescent wave, and n1Is the refractive index, n, of the objective lens 102 in TIRF illumination prior to the sample 1013 being imaged2For folding of the sample 1013 to be imagedThe refractive index. According to the formula, the larger the incident angle is, the smaller the transmission depth of the evanescent wave is, that is, the light intensity is performed according to the relation between the illumination depth and the brightness of the evanescent wave.
A method of depth imaging of TIRF illumination further comprising: intensity compensation is performed for evanescent wave illumination at different depths to correct for imaging intensity variations due to evanescent wave depth. As shown in fig. 3, the intensity of the different depths and the depth of illumination are inversely exponential.
The TIRF illumination depth imaging system applicable to the TIRF illumination depth imaging method comprises the following steps: a STORM imaging unit and a TIRF illumination unit; the TIRF illuminating units respectively generate evanescent waves with different transmission depths to irradiate the sample 1013 to be imaged, and the STORM imaging units image the sample 1013 to be imaged; the sample 1013 to be imaged is clamped between an upper slide 1011 and a lower slide 1012, immersion oil is filled between the lower slide 1012 and the objective lens 102, and the refractive indexes of the lower slide 1012, the immersion oil and the objective lens 102 are the same and are all larger than the refractive index of the sample 1013 to be imaged.
In this example, the refractive index n of the immersion oil1Refractive index n of sample 1013 to be imaged21.38; the total internal emission angle and the maximum aperture angle are illustrated in fig. 4, for example.
The TIRF lighting unit comprises: an illumination light source 104, an intermediate turning mirror 103, and an objective lens 102; the laser emitted by the illumination light source 104 is reflected by the intermediate fold-back mirror 103, transmitted by the objective lens 102, irradiated onto the lower slide 1012 for total reflection, and generates evanescent waves to irradiate onto the sample 1013 to be imaged. Further, the illumination light source 104 includes: a fiber optic light source 10421 and a collimating lens group 10411; the laser light emitted from the fiber light source 10421 passes through the collimating lens group 10411 and then irradiates the intermediate turning mirror 103.
The TIRF illumination units generating evanescent waves of different transmission depths respectively include: the incident angle on the sample 1013 to be imaged is changed by changing the movable members of the TIRF illumination unit, and a curve of the relationship between the moving position and the incident angle is obtained by measuring the relationship between the moving position of the movable members and the incident angle on the sample 1013 to be imaged at that time.
The TIRF illumination units respectively generate evanescent waves with different transmission depths to irradiate the sample 1013 to be imaged, and the STORM imaging units each image the sample 1013 to be imaged, including:
the method comprises the following steps: moving the movable piece to a position L0, wherein the depth of the evanescent wave is d0, the STORM imaging camera focuses to d0 for clear imaging, and the sample 1013 to be imaged is imaged as S0；
Step two: moving the movable piece to L1, wherein the corresponding depth of the evanescent wave is d0 +. DELTA.d, the depth of the evanescent wave is d1 ═ d0 +. DELTA.d, and the imaging of the sample 1013 to be imaged is S1(ii) a Wherein, the delta d is an equal division interval after N equal division is carried out on the transmission depth of the evanescent wave;
step three: moving the movable part to L2, wherein the corresponding depth of the evanescent wave is d0+2 delta d, moving the objective lens 102 in front of the sample 1013 to be imaged by a distance of 2 delta d/n1, wherein the objective lens 102 is focused at L2, the depth of the evanescent wave is d 2-d 0+2 delta d, and the image of the sample 1013 to be imaged is S2；
Step four: repeating the second step and the third step, and moving the movable piece until the position of the movable piece is LNAt this time, the sample 1013 to be imaged is imaged as SN(ii) a In the above steps, the objective lens 102 at the same position images two different evanescent wave depths each time;
step five: two adjacent samples 1013 to be imaged are sequentially subtracted to obtain Δ S1, Δ S2 and … Δ SN, and the subtracted images are images of the samples 1013 to be imaged at different depths.
When the Δ d is smaller, the objective lens 102 at the same position can be properly selected to image m different evanescent wave depths, that is, after only m moving parts move, the position of the objective lens 102 is adjusted again to image the next m moving parts. The steps are special examples, and m is 2;
the movable member includes: any one of moving the collimating lens group 10411, moving the fiber optic light source 10421, and moving the illumination light source 104. Fig. 5 shows that moving the collimating lens group 10411 changes the incidence angle of TIRF. FIG. 6 shows that moving fiber optic source 10421 changes the incidence angle of the TIRF. As shown in FIG. 7, moving the illumination source 104 changes the relative angle of the collimating lens group 10411 and the overall fiber to change the angle of incidence of the TIRF.
The above-mentioned embodiments are preferred embodiments of the present invention, and the present invention is not limited thereto, and any other modifications or equivalent substitutions that do not depart from the technical spirit of the present invention are included in the scope of the present invention.
Claims (10)
1. A method of depth imaging using TIRF illumination, comprising:
during TIRF illumination, incident light is totally reflected in front of a sample to be imaged, an evanescent wave is generated, and the generated evanescent wave is transmitted to the sample to be imaged; changing the incident angle of incident light to change the transmission depth of evanescent waves; when evanescent waves with different transmission depths respectively irradiate the samples to be imaged, the STORM imaging unit images the samples to be imaged, all the images of the samples to be imaged are calculated through a preset image reconstruction algorithm, the images of the samples to be imaged with different depths are obtained, and depth imaging is achieved.
2. The TIRF illuminated depth imaging method of claim 1, comprising:
s1, the incident angle of the incident light is theta during TIRF illumination1Transmission depth of H1Fluorescence imaging by STORM as S1；
S2, changing the incident angle to theta2At the time of the depth of transmission of H2Fluorescence imaging by STORM as S2(ii) a Wherein H2>H1；
S3, calculating the transmission depth H1-H2The corresponding image of the sample to be imaged is S2-S1；
S4, repeatedly changing different incidence angles until obtaining the transmission depth Hn-Hn-1Corresponding sample image S to be imagedn-Sn-1And imaging of samples to be imaged at different depths is realized.
3. The TIRF illuminated depth imaging method of claim 1, wherein the incident angle of the incident light is expressed in relation to the transmission depth of the evanescent wave as:
where θ is the incident angle of the incident light, d is the transmission depth of the evanescent wave, and n1Is the refractive index, n, of the objective lens in front of the sample to be imaged in TIRF illumination2Is the refractive index of the sample to be imaged.
4. The TIRF illuminated depth imaging method of claim 1, further comprising: intensity compensation is performed for evanescent wave illumination at different depths to correct for imaging intensity variations due to evanescent wave depth.
5. A TIRF illuminated depth imaging system, comprising: a STORM imaging unit and a TIRF illumination unit; the TIRF illuminating units respectively generate evanescent waves with different transmission depths to irradiate the samples to be imaged, and the STORM imaging units all image the samples to be imaged; the sample to be imaged is clamped between an upper glass slide and a lower glass slide, immersion oil is filled between the lower glass slide and the objective lens, and the refractive indexes of the lower glass slide, the immersion oil and the objective lens are the same and are all larger than the refractive index of the sample to be imaged.
6. The TIRF illuminated depth imaging system of claim 5, wherein the TIRF illumination unit comprises: an illumination light source, a middle fold-back mirror and an objective lens; laser emitted by the illumination light source is reflected by the intermediate turning mirror, is transmitted by the objective lens and then irradiates the lower glass slide for total reflection, and evanescent waves are generated and irradiate the sample to be imaged.
7. The TIRF illuminated depth imaging system of claim 6, wherein the illumination source comprises: a fiber light source and a collimating lens group; laser emitted by the optical fiber light source passes through the collimating lens group and then irradiates the middle turning mirror.
8. The TIRF illuminated depth imaging system of claim 7, wherein the TIRF illumination unit generating evanescent waves of different transmission depths, respectively, comprises:
the incidence angle of the sample to be imaged is changed by changing the movable piece of the TIRF lighting unit, and a relation curve of the moving position and the incidence angle is obtained by measuring the relation between the moving position of the movable piece and the incidence angle of the sample to be imaged at the moment.
9. The TIRF illuminated depth imaging system of claim 8, wherein the TIRF illumination units each generate evanescent waves of different transmission depths for impinging on the sample to be imaged, the STORM imaging units each imaging the sample to be imaged comprising:
the method comprises the following steps: moving the movable piece to a position L0, wherein the depth of the evanescent wave is d0, the STORM imaging camera focuses to d0 for clear imaging, and the imaging of the sample to be imaged is S0；
Step two: moving the movable piece to L1, wherein the corresponding depth of the evanescent wave is d0 +. DELTA.d, the depth of the evanescent wave is d1 ═ d0 +. DELTA.d, and the sample to be imaged is imaged as S1(ii) a Wherein, the delta d is an equal division interval after N equal division is carried out on the transmission depth of the evanescent wave;
step three: moving the movable piece to L2, wherein the corresponding depth of evanescent wave is d0+2 Δ d, and the sample to be imaged is imaged as S3；
Step four: continuously moving the movable piece until the position of the movable piece is LmMoving the objective lens in front of the sample to be imaged by a distance m delta d/n1 when the depth of the evanescent wave is d0+ m delta d, wherein the depth of the evanescent wave is d0+ m delta d, and the sample to be imaged is imaged as Sm；
Step five: repeating the second step to the fourth step, moving the movable piece until the position of the movable piece is LNWhen the sample to be imaged is imaged as SN；
Step six: and sequentially subtracting the images of two adjacent samples to be imaged to obtain delta S1, delta S2 and … delta SN respectively, wherein the images obtained by subtracting are the images of the samples to be imaged at different depths.
10. The TIRF illuminated depth imaging system of claim 9, wherein the movable member comprises: any one of a collimating lens group, a fiber optic light source, and an illumination light source.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011637113.4A CN112816410A (en) | 2020-12-31 | 2020-12-31 | Depth imaging method and system of TIRF illumination |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011637113.4A CN112816410A (en) | 2020-12-31 | 2020-12-31 | Depth imaging method and system of TIRF illumination |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN112816410A true CN112816410A (en) | 2021-05-18 |
Family
ID=75857349
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202011637113.4A Pending CN112816410A (en) | 2020-12-31 | 2020-12-31 | Depth imaging method and system of TIRF illumination |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN112816410A (en) |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000070329A1 (en) * | 1999-05-14 | 2000-11-23 | Brandeis University | Nucleic acid-based detection |
| CN1588003A (en) * | 2004-08-27 | 2005-03-02 | 中国科学院上海光学精密机械研究所 | Method for rapidly and longitudinally super-resolving single-molecule fluorescent sample |
| US20080151226A1 (en) * | 2003-09-25 | 2008-06-26 | Leica Microsystems Cms Gmbh | Method for analyzing a sample and microscope for evanescently illuminating the sample |
| CN103048272A (en) * | 2013-01-08 | 2013-04-17 | 浙江大学 | Frequency-shift super-resolution microimaging method and device based on evanescent field illumination |
| CN105807412A (en) * | 2016-04-07 | 2016-07-27 | 浙江大学 | Total internal reflection microscopy method and device based on free-form surface shaping |
-
2020
- 2020-12-31 CN CN202011637113.4A patent/CN112816410A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000070329A1 (en) * | 1999-05-14 | 2000-11-23 | Brandeis University | Nucleic acid-based detection |
| US20080151226A1 (en) * | 2003-09-25 | 2008-06-26 | Leica Microsystems Cms Gmbh | Method for analyzing a sample and microscope for evanescently illuminating the sample |
| CN1588003A (en) * | 2004-08-27 | 2005-03-02 | 中国科学院上海光学精密机械研究所 | Method for rapidly and longitudinally super-resolving single-molecule fluorescent sample |
| CN103048272A (en) * | 2013-01-08 | 2013-04-17 | 浙江大学 | Frequency-shift super-resolution microimaging method and device based on evanescent field illumination |
| CN105807412A (en) * | 2016-04-07 | 2016-07-27 | 浙江大学 | Total internal reflection microscopy method and device based on free-form surface shaping |
Non-Patent Citations (1)
| Title |
|---|
| 杨建宇 等: ""随机光学重构显微术及其应用研究进展"", 《红外与激光工程》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US6839469B2 (en) | Multiparallel three dimensional optical microscopy system | |
| US10116855B2 (en) | Autofocus method for microscope and microscope comprising autofocus device | |
| US9804377B2 (en) | Low numerical aperture exclusion imaging | |
| US9488818B2 (en) | Immersion objective for microscopes and use thereof | |
| WO2009153919A1 (en) | Microscope device and microscope device control program | |
| EP1941313A2 (en) | An optical system for illumination of an evanescent field | |
| CN103135220A (en) | Illuminating system for microscope and corresponding method | |
| CN111580261B (en) | Micro-imaging device based on epi-illumination | |
| JP5107003B2 (en) | Evanescent wave generator and observation apparatus using the same | |
| CN108387562B (en) | Method for adjusting axial position of pinhole in confocal microscope system | |
| JP2021516329A (en) | Lighting for fluorescence imaging using an objective lens | |
| CN111175954B (en) | Quick high-contrast image scanning microscopic imaging device based on Nipkow disk | |
| CN112816410A (en) | Depth imaging method and system of TIRF illumination | |
| CN110664369B (en) | Self-adaptive confocal line scanning harmonic microscopic imaging method and device | |
| US6940641B2 (en) | Fluorescence observation apparatus | |
| JP2018128580A (en) | Illumination device and microscope device | |
| CN214384623U (en) | High-resolution imaging system | |
| CN1715870A (en) | Phase object scan imaging method and its treating device | |
| CN210005784U (en) | digital pathological imaging equipment | |
| CN214409436U (en) | TIRF lighting system | |
| US20130321907A1 (en) | Adjustable Total Internal Reflectance Microscopy (TIRFM) Illuminator Apparatus | |
| CN117233947B (en) | Microscope illumination system, control method and microscopic imaging detection system | |
| Xue et al. | PN-ImTLSM facilitates high-throughput low background single-molecule localization microscopy deep in the cell | |
| US20220206279A1 (en) | Microscopy illumination apparatus, methods, and applications | |
| CN115656123A (en) | Automatic focusing device suitable for fluorescent microscope |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| CB02 | Change of applicant information |
Address after: No. 388, Lianyun Road, Huangpu District, Guangzhou, Guangdong 510530 Applicant after: Huangpu Material Research Institute Dawan District Guangdong Hong Kong and Macao Address before: No. 388, Lianyun Road, Huangpu District, Guangzhou, Guangdong 510530 Applicant before: Huangpu Institute of advanced materials, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences |
|
| CB02 | Change of applicant information | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210518 |
|
| RJ01 | Rejection of invention patent application after publication |